Western Blot eHandbook

View our step-by-step guide to western blotting including troubleshooting tips.

Step-by-Step Guide to Western Blotting

Understand the principles behind this common technique and learn how to achieve reliable and reproducible results.

Whether you are a student/researcher new to western blotting, or a skilled scientist, you’ll gain valuable insights to help you to master the technique.

You'll learn:

  • Main principles of western blotting
  • Troubleshooting tips
  • How to accurately analyze data
  • Proper controls, antibody validation, and data normalization
  • Fully automated Simple Western™ platforms from ProteinSimple, a Bio-Techne brand

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Western Blot Tip

Problem: High Uniform Background

Possible Reason & Solutions: Non-specific binding due to high antibody concentration

  • Lower concentration of primary or secondary antibody (titrations may be helpful).
  • Use a strong blocking agent in antibody diluents (milk is typically a stronger blocking agent than BSA).
  • Confirm that the background is not due to issues with the secondary antibody by omitting the primary antibody.